The cellulase cocktail produced by marine Aspergillus niger exhibits a property of salt-tolerance,which is of great potential in cellulose degradation in high salt environment.In order to explain the mechanism on the ...The cellulase cocktail produced by marine Aspergillus niger exhibits a property of salt-tolerance,which is of great potential in cellulose degradation in high salt environment.In order to explain the mechanism on the salttolerance of the cellulase cocktail produced by marine A.niger,six cellulase components(AnCel6,AnCel7A,AnCel7B,AnEGL,AnBGL1 and AnBGL2)were obtained by directed expression.Studies on their enzymatic properties revealed that oneβ-glucosidase(AnBGL2)and one endoglucanase(AnEGL)exhibited an outstanding salttolerant property,and one cellobiohydrolase(AnCel7B)exhibited a certain salt-tolerant property.Subsequent study revealed that the salt-tolerant An EGL and AnCel7B endowed the cellulase cocktail with stronger salttolerant property,while the salt-tolerant An BGL2 had no positive effect.Moreover,after overexpression of AnCel6,AnCel7A,AnCel7B and AnEGL,the activity of cellulase cocktail increased by 80%,70%,63%and 68%,respectively.However,the activity of cellulase cocktail was not improved after overexpression of AnBGL1 and AnBGL2.After mixed-strain fermentation with cellobiohydrolase recombinants(cel6 a,cel7a and cel7b recombinants)and endoglucanase recombinant(egl recombinant),the the activity of cellulase cocktail increased by 114%,102%and91%,respectively.展开更多
This study assessed the effects of lactic acid bacteria(LAB), cellulase, cellulase-producing Bacillus pumilus and their combinations on the fermentation characteristics, chemical composition, bacterial community and i...This study assessed the effects of lactic acid bacteria(LAB), cellulase, cellulase-producing Bacillus pumilus and their combinations on the fermentation characteristics, chemical composition, bacterial community and in vitro digestibility of alfalfa silage. A completely randomized design involving a 8(silage additives)×3 or 2(silage days) factorial arrangement of treatments was adopted in the present study. The 8 silage additive treatments were: untreated alfalfa(control); two commercial additives(GFJ and Chikuso-1); an originally selected LAB(Lactobacillus plantarum a214) isolated from alfalfa silage; a cellulase-producing Bacillus(CB) isolated from fresh alfalfa; cellulase(C); and the combined additives(a214+C and a214+CB). Silage fermentation characteristics, chemical composition and microorganism populations were analysed after 30, 60 and 65 days(60 days followed by exposure to air for five additional days). In vitro digestibility was analysed for 30 and 60 days. Compared with the other treatments, selected LAB a214, a214 combined with either C or CB, and Chikuso-1 had the decreased(P<0.001) pH and increased(P<0.001) lactic acid concentrations during the ensiling process, and there were no differences(P>0.05) among them. Fiber degradation was not significant(P≥0.054) in any C or CB treatments. The a214 treatment showed the highest(P=0.009) in vitro digestibility of dry matter(595.0 g kg^(–1) DM) after ensiling and the highest abundance of Lactobacillus(69.42 and 79.81%, respectively) on days 60 and 65, compared to all of other treatments. Overall, the silage quality of alfalfa was improved with the addition of a214, which indicates its potential as an alfalfa silage inoculant.展开更多
The object of this study was to determine the effect of cellulase and lactic acid bacteria (LAB) on fermentation quality and chemical composition of wheat straw silage. Silages were prepared using a small-scale fermen...The object of this study was to determine the effect of cellulase and lactic acid bacteria (LAB) on fermentation quality and chemical composition of wheat straw silage. Silages were prepared using a small-scale fermentation system and the moisture level was adjusted to 60% of fresh matter (FM) with deionized water. Treatments were designed as: control silage without additives, LAB inoculant Lactobacillus casei Z3-1 (1.0 × 106 cfu·g-1 of FM), commercial inoculant L. plantarum FG 1 (1.0 × 106 cfu·g-1 of FM), Z3-1 + cellulase and FG 1 + cellulase. The neutral detergent fiber (NDF), acid detergent fiber (ADF) and crude protein (CP) contents of the wheat straw prior to ensiling were 76.93%, 48.52% and 4.63% of dry matter (DM), respectively. After 30 days of fermentation, the silages treated with LAB and LAB + cellulase had a lower (P < 0.05) pH and higher (P < 0.05) lactic acid content than the control, and the coliform bacteria, yeast and mold were inhibited at the early stage of fermentation. Besides, silages treated with cellulase had lower (P < 0.05) values of ADF and NDF than the control. The results confirmed that the addition of cellulase and LAB contributed to improving the fermentation quality of wheat straw silage.展开更多
We investigated the effects of ultrasound treatment on cellulase adsorption and lignocellulose hydrolysis.The activity of cellulase remained constant upon lowpower ultrasound treatment(<120 W) and decreased using h...We investigated the effects of ultrasound treatment on cellulase adsorption and lignocellulose hydrolysis.The activity of cellulase remained constant upon lowpower ultrasound treatment(<120 W) and decreased using high-power ultrasound(>280 W).Oscillating cellulase adsorption occurred upon ultrasound treatment with any intensity.The maxima for desorption and adsorption were41.9 and 83.1%,respectively,during 1 h of 90 W ultrasound treatment at 50 °C.A comparison between the shorttime with long-time ultrasound experiments indicated that ultrasound treatment tended to desorb cellulase from substrate.However,ultrasound treatment also led to further surface erosion of biomass,which increased cellulase accessibility.These joint actions of ultrasound treatment induced the oscillating adsorption of cellulase.The increase in cellulase accessibility caused by ultrasound treatment led to a significant enhancement in lignocellulose hydrolysis.展开更多
This paper reports the purification and characterization of kinetic parameters of cellulase produced from Trichoderma viride under still culture solid state fermentation technique using cheap and an easily available a...This paper reports the purification and characterization of kinetic parameters of cellulase produced from Trichoderma viride under still culture solid state fermentation technique using cheap and an easily available agricultural waste material, wheat straw as growth supported substrate. Trichoderma viride was cultured in fermentation medium of wheat straw under some previously optimized growth conditions and maximum activity of 398±2.43U/mL obtained after stipulated fermentation time period. Cellulase was purified 2.33 fold with specific activity of 105U/mg in comparison to crude enzyme extract using ammonium sulfate precipitation, dialysis and Sephadex-G-100 column chromatography. The enzyme was shown to have a relative low molecular weight of 58kDa by sodium dodecyl sulphate poly-acrylamide gel electrophoresis. The purified enzyme displayed 6.5 and 55oC as an optimum pH and temperature respectively. Using carboxymethyl cellulose as substrate, the enzyme showed maximum activity (Vmax) of 148U/mL with its corresponding KM value of 68μM. Among activators/inhibitors SDS, EDTA, and Hg2+ showed inhibitory effect on purified cellulase whereas, the enzyme activated by Co2+ and Mn2+ at a concentration of 1mM. The purified cellulase was compatible with four local detergent brands with up to 20 days of shelf life at room temperature suggesting its potential as a detergent additive for improved washing therefore, it is concluded that it may be potentially useful for industrial purposes especially for detergent and laundry industry.展开更多
Cellulase is a complex enzyme that can decompose cellulose into glucose,and it could effectively treat cellulose waste. In this paper,it aims to explore development status and research progress of cellulase,and introd...Cellulase is a complex enzyme that can decompose cellulose into glucose,and it could effectively treat cellulose waste. In this paper,it aims to explore development status and research progress of cellulase,and introduce concept and action mechanism of cellulase,research situation of cellulase in molecular aspect,application of cellulase,and development of cellulase is also prospected.展开更多
A natural bacterial strain identified as Bacillus amyloliquefaciens MBAA3 using 16S rDNA partial genome sequencing has been studied for optimization of cellulase production.Statistical screening of media components fo...A natural bacterial strain identified as Bacillus amyloliquefaciens MBAA3 using 16S rDNA partial genome sequencing has been studied for optimization of cellulase production.Statistical screening of media components for production of cellulase by B.amyloliquefaciens MBAA3 was carried out by Plackett–Burman design.Plackett–Burman design showed CMC,MgSO4 and pH as significant components influencing the cellulase production from the media components screened by Plackett-Burman fractional factorial design.The optimum concentrations of these significant parameters were determined employing the response surface central composite design,involving three factors and five levels was adopted to acquire the best medium for the production of cellulase enzyme revealed concentration of CMC(1.84 g),MgSO4(0.275 g),and pH(8.5)in media for highest enzyme production.Response surface counter plots revealed that middle level of MgSO4 and middle level of CMC,higher level of CMC and lower level of pH and higher level of MgSO4 with lower level of pH increase the production of cellulase.After optimization cellulase activity increased by 6.81 fold.Presence of cellulase gene in MBAA3 was conformed by the amplification of genomic DNA of MBAA3.A PCR product of cellulase gene of 1500 bp was successfully amplified.The amplified gene was conformed by sequencing the amplified product and sequence was deposited in the gene bank under the accession number KF929416.展开更多
Microorganisms living in polar zones play an important part as the potential source of organic activity materials with low temperature characteristics in the biotechnological applications. A psychrotrophic bacterium (...Microorganisms living in polar zones play an important part as the potential source of organic activity materials with low temperature characteristics in the biotechnological applications. A psychrotrophic bacterium (strain Ar/w/b/75°/10/5), producing cellulase at low temperatures during late-exponential and early-stationary phases of cell growth, was isolated from sea ice-covered surface water in Chuckchi Sea, Arctic. This bacterium, with rod cells, was Gram-negative, slightly halophilic. Colony growing on agar plate was in black. Optimum growth temperature was 15℃. No cell growth was observed at 35℃ or above. Optimum salt concentration for cell growth was between 2 and 3 % of sodium chloride in media. Maximal cellulase activity was detected at a temperature of 35℃ and pH8. Cellulase was irreversibly inactivated when incubated at 55℃ within 30 min. Enzyme can be kept stable at the temperature no higher than 25℃. Of special interest was that this bacterium produced various extracellular enzymes including cellulase, amylase, agar hydrolase and protease, at low or moderate temperature conditions, which is certainly of it potential value for applications.展开更多
A cellulase producing bacterium (E3 strain) was isolated from fecal matter of elephant and identified as Bacillus sp. using 16S rDNA sequenced based molecular phylogenetic approach. While studying the effect of substr...A cellulase producing bacterium (E3 strain) was isolated from fecal matter of elephant and identified as Bacillus sp. using 16S rDNA sequenced based molecular phylogenetic approach. While studying the effect of substrates like Carboxymethyl cellulose (CMC), avicel, starch, maltose, sucrose, glucose, fructose, galactose and lactose on cellulase production, it was found that CMC was best carbon source induced cellulase production followed by lactose in this bacterial strain. A positive synergistic effect of lactose with CMC was also observed with enhancement of 5 - 6 times in cellulase production. The optimum cellulase production was recorded with 1% CMC and 1% lactose when added individually in the Omeliansky’s medium. The results showed that addition lactose with CMC greatly enhances the production and activity of various cellulase enzymes. The optimal fermentation conditions for the biosynthesis of cellulase by this strain were found to be temperature: 37℃, pH 7.0. The nitrogen source NH4Cl at 0.15% was optimum for cellulase production by this bacterium.展开更多
Lignocellulosic substrates are a good carbon source and provide rich growth media for a variety of microorganisms which prodLuce industrially important enzymes. Cellulases are a group of hydrolytic enzymes such as fil...Lignocellulosic substrates are a good carbon source and provide rich growth media for a variety of microorganisms which prodLuce industrially important enzymes. Cellulases are a group of hydrolytic enzymes such as filter paperase (FPase), carboxymethyl cellulase(CMCase) andβ-glucosidase-responsible for release of sugars in the bioconversion of the lignocellulosic biomass into a variety of value-added products. This study examined cellulase production by a newly isolated Aspergillus unguis on individual lignocellulosic substrates in solid state fermentation (SSF). The maximum peak production of enzymes varied from one substrate to another, however,based on the next best solid support and local availability of groundnut fodder supported maximum enzyme yields compared with other solid supports used in this study.Groundnut fodder supported significant production of FPase (5.9 FPU/g of substrate), CMCase (1.1 U/g of substrate) andβ-glucosidase activity (6.5 U/g of substrate) in SSF. Considerable secretion of protein (27.0 mg/g of substrate) on groundnut fodder was recorded. Constant increment of protein content in groundnut fodder due to cultivation of A. unguis is an interesting observation and it has implications for the improvement of nutritive value of groundnut fodder for cattle.展开更多
Central composite design (CCD)sp. JS14 in a solvent extraction was established with Response surface methodology (RSM). Solvent concentration, pH, temperature and retention time were selected as process variables to e...Central composite design (CCD)sp. JS14 in a solvent extraction was established with Response surface methodology (RSM). Solvent concentration, pH, temperature and retention time were selected as process variables to evaluate the purification impact factor in solvent precipitation, including the purification fold and % recovery. An experimental space with 13 purification fold and 23 recovery percentage recovery is achieved through the optimized condition based on the model. The molecular weight of the purified enzyme was estimated to be 32.5 KDa. Optimum activity of purified enzyme was at pH and temperature 6.5℃ and 40℃ respectively. Enzyme showed maximum activity with carboxymethyl cellulose as substrate with compare to rice husk, wheat straw and sucrose. The purified cellulase activity was inhibited by Na+, Cl- Mg2+ Tween 80 and EDTA.展开更多
The effects of various metal ions (Na+, K+, Ca2+, Mg2+, Al3+, Co2+) and anions (Cl–, SO2-4 and CH3COO–) on two cellulases were investigated. Fitting of the data to Michaelis-Menten kinetics showed that Al3+ noncom-p...The effects of various metal ions (Na+, K+, Ca2+, Mg2+, Al3+, Co2+) and anions (Cl–, SO2-4 and CH3COO–) on two cellulases were investigated. Fitting of the data to Michaelis-Menten kinetics showed that Al3+ noncom-petitively inhibited cellulase (Km = 22.68 g/L;Vmax = 0.269 mg/min at 5.0 mmol/L AlCl3) and Mg2+ competitively inhibited cellulase (Km = 50.0 g/L;Vmax = 0.434 mg/min at 10.0 mmol/L MgCl2) Different metal ions increased or decreased inhibition of cellulase activity slightly below 1 mmol/L and strongly over 10 mmol/L. The results indicated that filter paper activeity (FPA) was suitable for analysis of enzymatic saccharification with various lignocellulosic substrates whereas crude cellulase was suitable for applications in the biomass industry. Some metal ions were proved to inhibit cellulase reversibly.展开更多
Cellulose is the main structural component of lignocellulosic wastes that can be converted to sugars and biofuels by cellulase.Due to wide applications of this enzyme in various industries around the world,cellulase i...Cellulose is the main structural component of lignocellulosic wastes that can be converted to sugars and biofuels by cellulase.Due to wide applications of this enzyme in various industries around the world,cellulase is considered as the third industrial enzyme.The ability of thermophilic bacteria in the production of heat-stable cellulases has made them valuable tools in biotechnology.The aim of this study was isolation and molecular identification of cellulolytic thermophile bacteria from Dig Rostam hot spring and investigating their cellulase activity.Samples were taken from water and sediments of this hot spring,and cellulolytic bacteria were enriched in media containing cellulose as the only carbon source.The bacteria were incubated at 60℃,and single colonies were then isolated on solid media.Congo red assay was used as a quick test for the qualitative screening of cellulase activity.According to these qualitative results,four colonies named CDB1,CDB2,CDB3,and CDB4 were isolated,and their growth curve and some other characteristics were determined by biochemical assays.Moreover,endoglucanase,exoglucanase,and FPase activities of the isolates were investigated quantitatively.Results indicated that CDB1 exhibited the highest endoglucanase(0.096 U/mL)and exoglucanase(0.156 U/mL)activities among other isolates.16S rDNA partial sequencing indicated that CDB1 had 99%similarity to the genus Anoxybacillus,and the other isolates showed the highest similarity to the genus Geobacillus.The cellulase gene of CDB1 isolate with the highest cellulase activity was also cloned,and its sequence is reported for the first time.Further studies on this thermophilic enzyme might be useful for industrial applications.展开更多
Effects of dilute acid and acid steam pretreatments were inspected for cellulose production of Eucalyptus leaves through Box-Behenken design,a three variable factors for response surface methodology by Bacillus subtil...Effects of dilute acid and acid steam pretreatments were inspected for cellulose production of Eucalyptus leaves through Box-Behenken design,a three variable factors for response surface methodology by Bacillus subtilus K^(-1)8.Maximum cellulose production performed in 250 mL erlenmeyer flask with submerged fermentation attained at 50℃,p H 5,140 r·min^(-1) for 24 h.Results showed the efficient cellulose production from acid steam pretreatment(being autoclaved at 15 Psi for 15 min)than acid pretreatment.The optimum condition for maximum carboxymethyl cellulas(CMCase)was 1.811 IU·mL^(-1)·min^(-1)(0.8%acid conc.,10 g biomass loading,6 h reaction time)and filter paper activity(FPase)was 2.255 IU·mL^(-1)·min^(-1)(1%acid conc.,10 g biomass loading,8 h reaction time).Whereas,the acid steam maximum CMCase activity recorded was 2.585 IU·mL^(-1)·min^(-1)(0.8%acid conc.,15 g substrate loading and 8 h reaction time)and the highest FPase activity was 2.055 IU·mL^(-1)·min^(-1)(0.8%conc.,10 g biomass,6 h reaction time then autoclaved).Results revealed that acid pretreated Eucalyptus leaves were better lignocellulosic biomass for cellulose production by submerged fermentation.展开更多
1,2, 3,4 - butanetetracarboxylic acid (BTCA) offers an alternative to the conventional N-methlol compounds as a crosslinking agent for cellulose textiles. Cellulase treatment is an effective method to improve the hand...1,2, 3,4 - butanetetracarboxylic acid (BTCA) offers an alternative to the conventional N-methlol compounds as a crosslinking agent for cellulose textiles. Cellulase treatment is an effective method to improve the handle of the cotton fabric. Thus it is of particular interest to treat cotton fabric using cellulase and BTCA simultaneously.In this study, BTCA was applied to the cellulase pretreated cotton fabric, and softener was also used. The results show that the treated fabric does not only have good wrinkle-free property but also good handle.展开更多
The conversion of lignocellulosic biomass into biofuels or biochemicals typically involves a pretreatment process followed by the enzyme-catalyzed hydrolysis of cellulose and hemicellulose components to fermentable su...The conversion of lignocellulosic biomass into biofuels or biochemicals typically involves a pretreatment process followed by the enzyme-catalyzed hydrolysis of cellulose and hemicellulose components to fermentable sugars.Many factors can contribute to the recalcitrance of biomass,e.g.,the lignin content and structure,crystallinity of cellulose,degree of fiber polymerization,and hemicellulose content,among others.However,nonproductive binding between cellulase and lignin is the factor with the greatest impact on enzymatic hydrolysis.To reduce the nonproductive adsorption of enzymes on lignin and improve the efficiency of enzymatic hydrolysis,this review comprehensively summarized the progress that has been made in understanding the interactions between lignin and enzymes.Firstly,the effects of pretreatment techniques on lignin content and enzymatic hydrolysis were reviewed.The effects of lignin content and functional groups on enzymatic hydrolysis were then summarized.Methods for the preparation and characterization of lignin films were assessed.Finally,the methods applied to characterize the interactions between lignin and cellulase were reviewed,and methods for decreasing the nonproductive binding of enzymes to lignin were discussed.This review provides an overview of the current understanding of how lignin hinders the enzymatic hydrolysis of lignocellulosic biomass,and provides a theoretical basis for the development of more economical and effective methods and additives to reduce the interaction of lignin and enzymes to improve the efficiency of enzymatic hydrolysis.展开更多
This study evaluates the influence of growth parameters such as pH, temperature, Carboxy Methyl Cellulose (CMC) concentration and agitation on cellulase production from three bacterial strains, viz., Achromobacter xyl...This study evaluates the influence of growth parameters such as pH, temperature, Carboxy Methyl Cellulose (CMC) concentration and agitation on cellulase production from three bacterial strains, viz., Achromobacter xylosoxidans BSS4, Bacillus sp. BSS3 and Pseudomonas sp. BSS2 isolated from the wood-yards on Kallai river bank in Kerala. Production of cellulase by these isolates was detected using basal salt medium (BSM) with 0.5% CMC as supplement, and CMCase activity was confirmed by iodine test. Dinitrosalicylic acid method was employed for assaying the cellulase production by measuring the amount of glucose liberated in μmol/mL/min. Maximum enzyme production from Pseudomonas sp. BSS2 was at pH 8, 37℃ with 1% CMC and 150 rpm, and cellulase production increased from initial 49.84 U/mL to 91.28 U/mL after optimization. The highest enzyme activity from Bacillus sp. BSS3 was at pH 9, 37℃ with 1% CMC, 150 rpm, and cellulase production increased from initial 26.05 U/mL to 104.68 U/mL after optimization. The maximum enzyme production from A. xylosoxidans BSS4 was at pH 7, 40℃ with 0.5% CMC and 150 rpm, and cellulase production increased from initial 55.28 U/mL to 68.37 U/mL after optimization. Thus among the three isolates, Bacillus sp. BSS3 showed maximum enzyme yield which can be explored for further scale up studies with an industrial perspective.展开更多
Cellulase is an enzyme produced by fungi, bacteria, protozoa and termite, that hydrolyze cellulose. They are known for their diverse applications in industry and medicine. The aim of this study is to purify and invest...Cellulase is an enzyme produced by fungi, bacteria, protozoa and termite, that hydrolyze cellulose. They are known for their diverse applications in industry and medicine. The aim of this study is to purify and investigate cellulolytic properties of cellulase enzyme produced by Bacillus sphaericus CE-3 isolated from refuse dump in Nnamdi Azikiwe University, Awka, Nigeria. Enzyme was produced by submerged fermentation at 30°C for 30 h. The enzyme was purified to homogeneity by dialysis in 4M sucrose solution, ion-exchange chromatography on Q-Sepharose FF and by hydrophobic interaction chromatography on Phenyl Sepharose CL-4B. The relative molecular mass of the enzyme was estimated using SDS-Polyacrylamide gel electrophoresis. Effects of temperature, pH and metals on enzyme activity and stability and the relative rate of hydrolysis of various substrates were also studied. The Purification fold for the enzyme was 7.8, with 66.4 μ/mg specific activity protein and overall yield of 35.8. The relative molecular mass range of the enzyme was estimated between 22.3 kDa - 26.3 kDa. The enzyme was optimally active at pH 9.0 and 40°C, stable at pH 9.0 and unusually retained over 90% activity between 50°C - 100°C after 30 min incubation. It was strongly activated by Mn2+ but inhibited by Ba2+, Co2+, Hg2+, Pb2+, Cu2+, Sr2+, Fe2+, Ca2+ and Zn2+. The cellulase displayed high catalytic activity with untreated sawdust, followed by carboxymethyl cellulose, while sodium hydroxide treated sawdust was the least hydrolyzed. Since the enzyme is thermo-stable, alkalophilic and could utilize natural wastes like sawdust as substrate, it is obvious that it would be of great use in textile, starch processing and pulp and paper industries.展开更多
Today, demand exists for cost-effective production of industrially important enzymes from entire scientific sectors. By keeping in mind the extensive industrial applications of cellulase, this study was performed to i...Today, demand exists for cost-effective production of industrially important enzymes from entire scientific sectors. By keeping in mind the extensive industrial applications of cellulase, this study was performed to immobilize the indigenous enzyme produced from Trichoderma viride under pre-optimized SSF of an agricultural waste material, wheat straw. To enhance the bio-catalytic and tolerance properties of the present enzyme gel matrix immobilization engineering was applied. Previously, 2.33~fold purified novel cellulase was immobilized in to a xerogel matrix of TMOS and PTMS. FTIR spectroscopy confirmed the successful immobilization of cellulase. The free and immobilized cellulase was characterized and stability profile showed that after 24 h incubation, immobilization enhanced the thermo-stability up to 75% against 80℃ as compare to the free enzyme. Xerogel matrix immobilization enhanced the catalytic efficiency of entrapped enzyme than that of the free cellulase. Among activators/inhibitors SDS, EDTA, and Hg2+ showed inhibitory effect while, gel matrix immobilization enhanced 80% tolerance capacity of the cellulase against inactivating agents.展开更多
Industrial applications require enzymes highly stable and economically viable in terms of reusability. Enzyme immobilization is an exciting alternative to improve the stability of enzymatic processes. The immobilizati...Industrial applications require enzymes highly stable and economically viable in terms of reusability. Enzyme immobilization is an exciting alternative to improve the stability of enzymatic processes. The immobilization of two commercial enzymes is reported here (cellulase and xylanase) using three chemical methods (adsorption, reticulation, and crosslinking-adsorption) and two polymeric supports (alginate-chitin and chitosan-chitin). The optimal pH for binding was 4.5 for cellulase and 5.0 for xylanase, and the optimal enzyme concentrations were 170 μg/mL and 127.5 μg/mL respectively, being the chitosan and the ideal support. In some cases, a low concentration of crosslinking agent (glutaraldehyde) improved stability of the immobilization process. Biotechnological characterization showed that the reusability of enzymes was the most striking finding, particularly of immobilized cellulase using glutaraldehyde, which after 19 cycles retained 64% activity. These results confirm the economic and biotechnical advantages of enzyme immobilization for a range of industrial applications.展开更多
基金supported by National Natural Science Foundation of China(21576233,21878263)Fundamental Research Funds for the Central Universities。
文摘The cellulase cocktail produced by marine Aspergillus niger exhibits a property of salt-tolerance,which is of great potential in cellulose degradation in high salt environment.In order to explain the mechanism on the salttolerance of the cellulase cocktail produced by marine A.niger,six cellulase components(AnCel6,AnCel7A,AnCel7B,AnEGL,AnBGL1 and AnBGL2)were obtained by directed expression.Studies on their enzymatic properties revealed that oneβ-glucosidase(AnBGL2)and one endoglucanase(AnEGL)exhibited an outstanding salttolerant property,and one cellobiohydrolase(AnCel7B)exhibited a certain salt-tolerant property.Subsequent study revealed that the salt-tolerant An EGL and AnCel7B endowed the cellulase cocktail with stronger salttolerant property,while the salt-tolerant An BGL2 had no positive effect.Moreover,after overexpression of AnCel6,AnCel7A,AnCel7B and AnEGL,the activity of cellulase cocktail increased by 80%,70%,63%and 68%,respectively.However,the activity of cellulase cocktail was not improved after overexpression of AnBGL1 and AnBGL2.After mixed-strain fermentation with cellobiohydrolase recombinants(cel6 a,cel7a and cel7b recombinants)and endoglucanase recombinant(egl recombinant),the the activity of cellulase cocktail increased by 114%,102%and91%,respectively.
基金supported by the National Key R&D Program of China (2017YFD0502102)the National Technology Leader “Ten Thousand People Plan” of China (201502510410040)the National Key Technology R&D Program of China during the 12th Five-year Plan period of China (2011BAD17B02)
文摘This study assessed the effects of lactic acid bacteria(LAB), cellulase, cellulase-producing Bacillus pumilus and their combinations on the fermentation characteristics, chemical composition, bacterial community and in vitro digestibility of alfalfa silage. A completely randomized design involving a 8(silage additives)×3 or 2(silage days) factorial arrangement of treatments was adopted in the present study. The 8 silage additive treatments were: untreated alfalfa(control); two commercial additives(GFJ and Chikuso-1); an originally selected LAB(Lactobacillus plantarum a214) isolated from alfalfa silage; a cellulase-producing Bacillus(CB) isolated from fresh alfalfa; cellulase(C); and the combined additives(a214+C and a214+CB). Silage fermentation characteristics, chemical composition and microorganism populations were analysed after 30, 60 and 65 days(60 days followed by exposure to air for five additional days). In vitro digestibility was analysed for 30 and 60 days. Compared with the other treatments, selected LAB a214, a214 combined with either C or CB, and Chikuso-1 had the decreased(P<0.001) pH and increased(P<0.001) lactic acid concentrations during the ensiling process, and there were no differences(P>0.05) among them. Fiber degradation was not significant(P≥0.054) in any C or CB treatments. The a214 treatment showed the highest(P=0.009) in vitro digestibility of dry matter(595.0 g kg^(–1) DM) after ensiling and the highest abundance of Lactobacillus(69.42 and 79.81%, respectively) on days 60 and 65, compared to all of other treatments. Overall, the silage quality of alfalfa was improved with the addition of a214, which indicates its potential as an alfalfa silage inoculant.
文摘The object of this study was to determine the effect of cellulase and lactic acid bacteria (LAB) on fermentation quality and chemical composition of wheat straw silage. Silages were prepared using a small-scale fermentation system and the moisture level was adjusted to 60% of fresh matter (FM) with deionized water. Treatments were designed as: control silage without additives, LAB inoculant Lactobacillus casei Z3-1 (1.0 × 106 cfu·g-1 of FM), commercial inoculant L. plantarum FG 1 (1.0 × 106 cfu·g-1 of FM), Z3-1 + cellulase and FG 1 + cellulase. The neutral detergent fiber (NDF), acid detergent fiber (ADF) and crude protein (CP) contents of the wheat straw prior to ensiling were 76.93%, 48.52% and 4.63% of dry matter (DM), respectively. After 30 days of fermentation, the silages treated with LAB and LAB + cellulase had a lower (P < 0.05) pH and higher (P < 0.05) lactic acid content than the control, and the coliform bacteria, yeast and mold were inhibited at the early stage of fermentation. Besides, silages treated with cellulase had lower (P < 0.05) values of ADF and NDF than the control. The results confirmed that the addition of cellulase and LAB contributed to improving the fermentation quality of wheat straw silage.
基金Supported by the Natural Science Foundation of China (No.51473115 and No.21276192)the Research Project of Chongqing Education Commission (No.KJ1500632)
文摘We investigated the effects of ultrasound treatment on cellulase adsorption and lignocellulose hydrolysis.The activity of cellulase remained constant upon lowpower ultrasound treatment(<120 W) and decreased using high-power ultrasound(>280 W).Oscillating cellulase adsorption occurred upon ultrasound treatment with any intensity.The maxima for desorption and adsorption were41.9 and 83.1%,respectively,during 1 h of 90 W ultrasound treatment at 50 °C.A comparison between the shorttime with long-time ultrasound experiments indicated that ultrasound treatment tended to desorb cellulase from substrate.However,ultrasound treatment also led to further surface erosion of biomass,which increased cellulase accessibility.These joint actions of ultrasound treatment induced the oscillating adsorption of cellulase.The increase in cellulase accessibility caused by ultrasound treatment led to a significant enhancement in lignocellulose hydrolysis.
文摘This paper reports the purification and characterization of kinetic parameters of cellulase produced from Trichoderma viride under still culture solid state fermentation technique using cheap and an easily available agricultural waste material, wheat straw as growth supported substrate. Trichoderma viride was cultured in fermentation medium of wheat straw under some previously optimized growth conditions and maximum activity of 398±2.43U/mL obtained after stipulated fermentation time period. Cellulase was purified 2.33 fold with specific activity of 105U/mg in comparison to crude enzyme extract using ammonium sulfate precipitation, dialysis and Sephadex-G-100 column chromatography. The enzyme was shown to have a relative low molecular weight of 58kDa by sodium dodecyl sulphate poly-acrylamide gel electrophoresis. The purified enzyme displayed 6.5 and 55oC as an optimum pH and temperature respectively. Using carboxymethyl cellulose as substrate, the enzyme showed maximum activity (Vmax) of 148U/mL with its corresponding KM value of 68μM. Among activators/inhibitors SDS, EDTA, and Hg2+ showed inhibitory effect on purified cellulase whereas, the enzyme activated by Co2+ and Mn2+ at a concentration of 1mM. The purified cellulase was compatible with four local detergent brands with up to 20 days of shelf life at room temperature suggesting its potential as a detergent additive for improved washing therefore, it is concluded that it may be potentially useful for industrial purposes especially for detergent and laundry industry.
文摘Cellulase is a complex enzyme that can decompose cellulose into glucose,and it could effectively treat cellulose waste. In this paper,it aims to explore development status and research progress of cellulase,and introduce concept and action mechanism of cellulase,research situation of cellulase in molecular aspect,application of cellulase,and development of cellulase is also prospected.
文摘A natural bacterial strain identified as Bacillus amyloliquefaciens MBAA3 using 16S rDNA partial genome sequencing has been studied for optimization of cellulase production.Statistical screening of media components for production of cellulase by B.amyloliquefaciens MBAA3 was carried out by Plackett–Burman design.Plackett–Burman design showed CMC,MgSO4 and pH as significant components influencing the cellulase production from the media components screened by Plackett-Burman fractional factorial design.The optimum concentrations of these significant parameters were determined employing the response surface central composite design,involving three factors and five levels was adopted to acquire the best medium for the production of cellulase enzyme revealed concentration of CMC(1.84 g),MgSO4(0.275 g),and pH(8.5)in media for highest enzyme production.Response surface counter plots revealed that middle level of MgSO4 and middle level of CMC,higher level of CMC and lower level of pH and higher level of MgSO4 with lower level of pH increase the production of cellulase.After optimization cellulase activity increased by 6.81 fold.Presence of cellulase gene in MBAA3 was conformed by the amplification of genomic DNA of MBAA3.A PCR product of cellulase gene of 1500 bp was successfully amplified.The amplified gene was conformed by sequencing the amplified product and sequence was deposited in the gene bank under the accession number KF929416.
基金This work was supported in part by State Oceanic Administration(No.99614)the State Ministry of Science and Technology(No.2001 DIA50040 6)National Nature Science Foundation of China(No.40006010).
文摘Microorganisms living in polar zones play an important part as the potential source of organic activity materials with low temperature characteristics in the biotechnological applications. A psychrotrophic bacterium (strain Ar/w/b/75°/10/5), producing cellulase at low temperatures during late-exponential and early-stationary phases of cell growth, was isolated from sea ice-covered surface water in Chuckchi Sea, Arctic. This bacterium, with rod cells, was Gram-negative, slightly halophilic. Colony growing on agar plate was in black. Optimum growth temperature was 15℃. No cell growth was observed at 35℃ or above. Optimum salt concentration for cell growth was between 2 and 3 % of sodium chloride in media. Maximal cellulase activity was detected at a temperature of 35℃ and pH8. Cellulase was irreversibly inactivated when incubated at 55℃ within 30 min. Enzyme can be kept stable at the temperature no higher than 25℃. Of special interest was that this bacterium produced various extracellular enzymes including cellulase, amylase, agar hydrolase and protease, at low or moderate temperature conditions, which is certainly of it potential value for applications.
文摘A cellulase producing bacterium (E3 strain) was isolated from fecal matter of elephant and identified as Bacillus sp. using 16S rDNA sequenced based molecular phylogenetic approach. While studying the effect of substrates like Carboxymethyl cellulose (CMC), avicel, starch, maltose, sucrose, glucose, fructose, galactose and lactose on cellulase production, it was found that CMC was best carbon source induced cellulase production followed by lactose in this bacterial strain. A positive synergistic effect of lactose with CMC was also observed with enhancement of 5 - 6 times in cellulase production. The optimum cellulase production was recorded with 1% CMC and 1% lactose when added individually in the Omeliansky’s medium. The results showed that addition lactose with CMC greatly enhances the production and activity of various cellulase enzymes. The optimal fermentation conditions for the biosynthesis of cellulase by this strain were found to be temperature: 37℃, pH 7.0. The nitrogen source NH4Cl at 0.15% was optimum for cellulase production by this bacterium.
文摘Lignocellulosic substrates are a good carbon source and provide rich growth media for a variety of microorganisms which prodLuce industrially important enzymes. Cellulases are a group of hydrolytic enzymes such as filter paperase (FPase), carboxymethyl cellulase(CMCase) andβ-glucosidase-responsible for release of sugars in the bioconversion of the lignocellulosic biomass into a variety of value-added products. This study examined cellulase production by a newly isolated Aspergillus unguis on individual lignocellulosic substrates in solid state fermentation (SSF). The maximum peak production of enzymes varied from one substrate to another, however,based on the next best solid support and local availability of groundnut fodder supported maximum enzyme yields compared with other solid supports used in this study.Groundnut fodder supported significant production of FPase (5.9 FPU/g of substrate), CMCase (1.1 U/g of substrate) andβ-glucosidase activity (6.5 U/g of substrate) in SSF. Considerable secretion of protein (27.0 mg/g of substrate) on groundnut fodder was recorded. Constant increment of protein content in groundnut fodder due to cultivation of A. unguis is an interesting observation and it has implications for the improvement of nutritive value of groundnut fodder for cattle.
文摘Central composite design (CCD)sp. JS14 in a solvent extraction was established with Response surface methodology (RSM). Solvent concentration, pH, temperature and retention time were selected as process variables to evaluate the purification impact factor in solvent precipitation, including the purification fold and % recovery. An experimental space with 13 purification fold and 23 recovery percentage recovery is achieved through the optimized condition based on the model. The molecular weight of the purified enzyme was estimated to be 32.5 KDa. Optimum activity of purified enzyme was at pH and temperature 6.5℃ and 40℃ respectively. Enzyme showed maximum activity with carboxymethyl cellulose as substrate with compare to rice husk, wheat straw and sucrose. The purified cellulase activity was inhibited by Na+, Cl- Mg2+ Tween 80 and EDTA.
文摘The effects of various metal ions (Na+, K+, Ca2+, Mg2+, Al3+, Co2+) and anions (Cl–, SO2-4 and CH3COO–) on two cellulases were investigated. Fitting of the data to Michaelis-Menten kinetics showed that Al3+ noncom-petitively inhibited cellulase (Km = 22.68 g/L;Vmax = 0.269 mg/min at 5.0 mmol/L AlCl3) and Mg2+ competitively inhibited cellulase (Km = 50.0 g/L;Vmax = 0.434 mg/min at 10.0 mmol/L MgCl2) Different metal ions increased or decreased inhibition of cellulase activity slightly below 1 mmol/L and strongly over 10 mmol/L. The results indicated that filter paper activeity (FPA) was suitable for analysis of enzymatic saccharification with various lignocellulosic substrates whereas crude cellulase was suitable for applications in the biomass industry. Some metal ions were proved to inhibit cellulase reversibly.
基金a grant(3/22775)from Ferdowsi University of Mashhad.
文摘Cellulose is the main structural component of lignocellulosic wastes that can be converted to sugars and biofuels by cellulase.Due to wide applications of this enzyme in various industries around the world,cellulase is considered as the third industrial enzyme.The ability of thermophilic bacteria in the production of heat-stable cellulases has made them valuable tools in biotechnology.The aim of this study was isolation and molecular identification of cellulolytic thermophile bacteria from Dig Rostam hot spring and investigating their cellulase activity.Samples were taken from water and sediments of this hot spring,and cellulolytic bacteria were enriched in media containing cellulose as the only carbon source.The bacteria were incubated at 60℃,and single colonies were then isolated on solid media.Congo red assay was used as a quick test for the qualitative screening of cellulase activity.According to these qualitative results,four colonies named CDB1,CDB2,CDB3,and CDB4 were isolated,and their growth curve and some other characteristics were determined by biochemical assays.Moreover,endoglucanase,exoglucanase,and FPase activities of the isolates were investigated quantitatively.Results indicated that CDB1 exhibited the highest endoglucanase(0.096 U/mL)and exoglucanase(0.156 U/mL)activities among other isolates.16S rDNA partial sequencing indicated that CDB1 had 99%similarity to the genus Anoxybacillus,and the other isolates showed the highest similarity to the genus Geobacillus.The cellulase gene of CDB1 isolate with the highest cellulase activity was also cloned,and its sequence is reported for the first time.Further studies on this thermophilic enzyme might be useful for industrial applications.
文摘Effects of dilute acid and acid steam pretreatments were inspected for cellulose production of Eucalyptus leaves through Box-Behenken design,a three variable factors for response surface methodology by Bacillus subtilus K^(-1)8.Maximum cellulose production performed in 250 mL erlenmeyer flask with submerged fermentation attained at 50℃,p H 5,140 r·min^(-1) for 24 h.Results showed the efficient cellulose production from acid steam pretreatment(being autoclaved at 15 Psi for 15 min)than acid pretreatment.The optimum condition for maximum carboxymethyl cellulas(CMCase)was 1.811 IU·mL^(-1)·min^(-1)(0.8%acid conc.,10 g biomass loading,6 h reaction time)and filter paper activity(FPase)was 2.255 IU·mL^(-1)·min^(-1)(1%acid conc.,10 g biomass loading,8 h reaction time).Whereas,the acid steam maximum CMCase activity recorded was 2.585 IU·mL^(-1)·min^(-1)(0.8%acid conc.,15 g substrate loading and 8 h reaction time)and the highest FPase activity was 2.055 IU·mL^(-1)·min^(-1)(0.8%conc.,10 g biomass,6 h reaction time then autoclaved).Results revealed that acid pretreated Eucalyptus leaves were better lignocellulosic biomass for cellulose production by submerged fermentation.
文摘1,2, 3,4 - butanetetracarboxylic acid (BTCA) offers an alternative to the conventional N-methlol compounds as a crosslinking agent for cellulose textiles. Cellulase treatment is an effective method to improve the handle of the cotton fabric. Thus it is of particular interest to treat cotton fabric using cellulase and BTCA simultaneously.In this study, BTCA was applied to the cellulase pretreated cotton fabric, and softener was also used. The results show that the treated fabric does not only have good wrinkle-free property but also good handle.
基金financially supported by Innovation Project of Guangxi Graduate Education (YCBZ2019017)Guangxi Natural Science Fund (2018JJA130224)Guangxi Key Laboratory of Clean Pulping and Pollution Control Fund (ZR2018057)
文摘The conversion of lignocellulosic biomass into biofuels or biochemicals typically involves a pretreatment process followed by the enzyme-catalyzed hydrolysis of cellulose and hemicellulose components to fermentable sugars.Many factors can contribute to the recalcitrance of biomass,e.g.,the lignin content and structure,crystallinity of cellulose,degree of fiber polymerization,and hemicellulose content,among others.However,nonproductive binding between cellulase and lignin is the factor with the greatest impact on enzymatic hydrolysis.To reduce the nonproductive adsorption of enzymes on lignin and improve the efficiency of enzymatic hydrolysis,this review comprehensively summarized the progress that has been made in understanding the interactions between lignin and enzymes.Firstly,the effects of pretreatment techniques on lignin content and enzymatic hydrolysis were reviewed.The effects of lignin content and functional groups on enzymatic hydrolysis were then summarized.Methods for the preparation and characterization of lignin films were assessed.Finally,the methods applied to characterize the interactions between lignin and cellulase were reviewed,and methods for decreasing the nonproductive binding of enzymes to lignin were discussed.This review provides an overview of the current understanding of how lignin hinders the enzymatic hydrolysis of lignocellulosic biomass,and provides a theoretical basis for the development of more economical and effective methods and additives to reduce the interaction of lignin and enzymes to improve the efficiency of enzymatic hydrolysis.
文摘This study evaluates the influence of growth parameters such as pH, temperature, Carboxy Methyl Cellulose (CMC) concentration and agitation on cellulase production from three bacterial strains, viz., Achromobacter xylosoxidans BSS4, Bacillus sp. BSS3 and Pseudomonas sp. BSS2 isolated from the wood-yards on Kallai river bank in Kerala. Production of cellulase by these isolates was detected using basal salt medium (BSM) with 0.5% CMC as supplement, and CMCase activity was confirmed by iodine test. Dinitrosalicylic acid method was employed for assaying the cellulase production by measuring the amount of glucose liberated in μmol/mL/min. Maximum enzyme production from Pseudomonas sp. BSS2 was at pH 8, 37℃ with 1% CMC and 150 rpm, and cellulase production increased from initial 49.84 U/mL to 91.28 U/mL after optimization. The highest enzyme activity from Bacillus sp. BSS3 was at pH 9, 37℃ with 1% CMC, 150 rpm, and cellulase production increased from initial 26.05 U/mL to 104.68 U/mL after optimization. The maximum enzyme production from A. xylosoxidans BSS4 was at pH 7, 40℃ with 0.5% CMC and 150 rpm, and cellulase production increased from initial 55.28 U/mL to 68.37 U/mL after optimization. Thus among the three isolates, Bacillus sp. BSS3 showed maximum enzyme yield which can be explored for further scale up studies with an industrial perspective.
文摘Cellulase is an enzyme produced by fungi, bacteria, protozoa and termite, that hydrolyze cellulose. They are known for their diverse applications in industry and medicine. The aim of this study is to purify and investigate cellulolytic properties of cellulase enzyme produced by Bacillus sphaericus CE-3 isolated from refuse dump in Nnamdi Azikiwe University, Awka, Nigeria. Enzyme was produced by submerged fermentation at 30°C for 30 h. The enzyme was purified to homogeneity by dialysis in 4M sucrose solution, ion-exchange chromatography on Q-Sepharose FF and by hydrophobic interaction chromatography on Phenyl Sepharose CL-4B. The relative molecular mass of the enzyme was estimated using SDS-Polyacrylamide gel electrophoresis. Effects of temperature, pH and metals on enzyme activity and stability and the relative rate of hydrolysis of various substrates were also studied. The Purification fold for the enzyme was 7.8, with 66.4 μ/mg specific activity protein and overall yield of 35.8. The relative molecular mass range of the enzyme was estimated between 22.3 kDa - 26.3 kDa. The enzyme was optimally active at pH 9.0 and 40°C, stable at pH 9.0 and unusually retained over 90% activity between 50°C - 100°C after 30 min incubation. It was strongly activated by Mn2+ but inhibited by Ba2+, Co2+, Hg2+, Pb2+, Cu2+, Sr2+, Fe2+, Ca2+ and Zn2+. The cellulase displayed high catalytic activity with untreated sawdust, followed by carboxymethyl cellulose, while sodium hydroxide treated sawdust was the least hydrolyzed. Since the enzyme is thermo-stable, alkalophilic and could utilize natural wastes like sawdust as substrate, it is obvious that it would be of great use in textile, starch processing and pulp and paper industries.
文摘Today, demand exists for cost-effective production of industrially important enzymes from entire scientific sectors. By keeping in mind the extensive industrial applications of cellulase, this study was performed to immobilize the indigenous enzyme produced from Trichoderma viride under pre-optimized SSF of an agricultural waste material, wheat straw. To enhance the bio-catalytic and tolerance properties of the present enzyme gel matrix immobilization engineering was applied. Previously, 2.33~fold purified novel cellulase was immobilized in to a xerogel matrix of TMOS and PTMS. FTIR spectroscopy confirmed the successful immobilization of cellulase. The free and immobilized cellulase was characterized and stability profile showed that after 24 h incubation, immobilization enhanced the thermo-stability up to 75% against 80℃ as compare to the free enzyme. Xerogel matrix immobilization enhanced the catalytic efficiency of entrapped enzyme than that of the free cellulase. Among activators/inhibitors SDS, EDTA, and Hg2+ showed inhibitory effect while, gel matrix immobilization enhanced 80% tolerance capacity of the cellulase against inactivating agents.
文摘Industrial applications require enzymes highly stable and economically viable in terms of reusability. Enzyme immobilization is an exciting alternative to improve the stability of enzymatic processes. The immobilization of two commercial enzymes is reported here (cellulase and xylanase) using three chemical methods (adsorption, reticulation, and crosslinking-adsorption) and two polymeric supports (alginate-chitin and chitosan-chitin). The optimal pH for binding was 4.5 for cellulase and 5.0 for xylanase, and the optimal enzyme concentrations were 170 μg/mL and 127.5 μg/mL respectively, being the chitosan and the ideal support. In some cases, a low concentration of crosslinking agent (glutaraldehyde) improved stability of the immobilization process. Biotechnological characterization showed that the reusability of enzymes was the most striking finding, particularly of immobilized cellulase using glutaraldehyde, which after 19 cycles retained 64% activity. These results confirm the economic and biotechnical advantages of enzyme immobilization for a range of industrial applications.
